Several arenaviruses cause hemorrhagic fever (HF) disease in humans and pose a serious public health problem in their endemic regions. Thus, Lassa virus (LASV) infects several hundred thousand individuals yearly in West Africa resulting in a high number of Lassa fever (LF) cases associated with high morbidity and mortality. Notably, increased traveling has led to the importation of LF cases into non-endemic metropolitan areas around the globe. Besides their impact in public health, several arenaviruses, including LASV, have features that make them a credible biodefense threat. There are no FDA-licensed vaccines to combat arenavirus infections and current anti-arenaviral therapy is limited to the off-label use of ribavirin, which is only partially effective and associated with side effects. The significance of arenaviruses, including LASV, in human health and biodefense readiness, together with the limited existing armamentarium to combat their infections, highlight the importance of developing effective vaccines to combat arenavirus induced disease in humans. The central goal of this application is to test the novel hypothesis that a codon deoptimized (CD)-based strategy can be used for the development of a safe, immunogenic, stable and protective live-attenuated vaccines (LAV) to combat HF disease caused by LASV infection. Moreover, we also hypothesize that a recombinant LASV expressing a codon deoptimized (CD) glycoprotein (GP), rLASV/GPCD, will prove investigators with a valid LASV surrogate that could be safely used in BSL2 facilities to facilitate the investigation of LASV-host cell interactions as well as the identification and characterization of anti- LASV candidate therapeutics without the need of BSL4 containment, currently needed to work with live forms of this important human biodefense pathogen. The bases for our hypothesis are: 1) Existing evidence that CD can be used to generate attenuated viruses, including arenaviruses, that are able to provide protection against a challenge with wild-type (WT) virus. 2) CD LAV have identical immunogenic properties than WT virus, but the large number (>100s) of silent mutations within the genome of CD LAV makes impossible its reversion to a virulent strain. 3) CD LAV, including arenaviruses, are able to grow to high titers, compatible with vaccine production, in FDA-approved Vero cells for vaccine production. 4) Generation of CD recombinant LASV can be rapidly achieved using our state-of-the-art reverse genetics technologies. To test our hypothesis, we will first use the prototypic LCMV system that provides us with an excellent reference virus that can be safely used in BSL2 facilities to test whether a CD LASV GP can convert a virulent strain into an attenuated (Aim 1) and stable (Aim 2) form still able to induce protective immunity (Aim 3) using the well characterized mouse model of LCMV infection. We will use this knowledge to generate a rLASV/GPCD whose potential as LAV will be evaluated using the well-established guinea pig model of LF disease (Aim 4).